The proposed studies focus on the mechanisms that underly the functions of the blood-retinal barrier.
The specific aims are: nature and mode of formation of luminal and abluminal endothelial vesicles, the mechanism of receptor-mediated endocytosis in vascular endothelial cells, the role of plasma membrane-associated phosphatases in active transport as it relates to permeable vessels, and the role of vesicular transport in breakdown of the blood-retinal barrier. Also included are related studies of antigenic components in the blood vessel basal lamina and study of the cytoskeleton in endothelial cells. These various studies are aimed at gaining a better understanding of retinal vascular function in normal and abnormal conditions. Luminal and abluminal endothelial vesicles will be studied in rapidly frozen, freeze-substituted tissues to verify the morphology of the vesicles and to determine the ratio of free (endocytic) to attached vesicles. Two physiologically important molecules, heparin and insulin, will be used to examine the pathways of receptor mediated endocytosis. Animals will be perfused with gold-ligand conjugates and examined for evidence of endocytic transport. We also plan to compare the localization of Na+,K+ ATPase, alkaline phosphatase and 5' nucleotidase activities in normal blood vessels and in those of streptozotocin-diabetic rats and galactose-fed rats which exhibit a diabetic-like condition. Breakdown of the blood-retinal barrier at the level of retinal capillaries will be induced by perfusion of animals with heparatinase, an enzyme which degrades heparan sulfate, and which has been shown to cause capillary leakage. This system will be used to investigate the morphology of luminal vesicles in frozen tissue and Possible changes in the polarity of the phosphatases mentioned above. In related work, several antigens, including type IV collagen, laminin, fibronectin and heparan sulfate proteoglycan will be localized in the vascular basal lamina using post- embedding, immunogold methods. The localizations obtained for normal vessels will be compared with those found in streptozotocin- diabetic and galactose-fed rats both of which show thickening of the vascular basal lamina. Results will be quantified using morphometric analysis. Finally, the endothelial cytoskeleton will be examined in normal arteriolar endothelial cells and in those of spontaneously, hypertensive-stroke prone rats.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
2R01EY004831-06
Application #
3259363
Study Section
Visual Sciences A Study Section (VISA)
Project Start
1983-09-01
Project End
1993-09-29
Budget Start
1988-09-30
Budget End
1989-09-29
Support Year
6
Fiscal Year
1988
Total Cost
Indirect Cost
Name
Wayne State University
Department
Type
Schools of Medicine
DUNS #
City
Detroit
State
MI
Country
United States
Zip Code
48202
Wang, J; Essner, E; Shichi, H (1995) Tissue repair in the iris in experimental autoimmune uveoretinitis. Exp Mol Pathol 63:41-51
Wang, J; Essner, E; Shichi, H (1994) Ultrastructural and immunocytochemical studies of smooth muscle cells in iris arterioles of rats with experimental autoimmune uveoretinitis. Exp Mol Pathol 61:153-63
Wang, J; Lin, W L; Essner, E et al. (1994) Ultrastructural and immunocytochemical studies of iris vessels in rats with experimental autoimmune uveoretinitis. Curr Eye Res 13:747-54
Lin, W L; Essner, E; McCarthy, K J et al. (1992) Ultrastructural immunocytochemical localization of chondroitin sulfate proteoglycan in Bruch's membrane of the rat. Invest Ophthalmol Vis Sci 33:2072-5
Mahalak, S M; Lin, W L; Essner, E et al. (1991) Increased immunoreactivity of collagen types I, III and V, fibronectin and TGF-beta in retinal vessels of rats with experimental autoimmune uveoretinitis. Curr Eye Res 10:1059-63
Shichi, H; Mahalak, S M; Sakamoto, S et al. (1991) Immunocytochemical localization of gamma-glutamyl transpeptidase in porcine ciliary epithelium. Exp Eye Res 53:39-46
Lin, W L; Essner, E; Shichi, H (1991) Breakdown of the blood-retinal barrier in S-antigen-induced uveoretinitis in rats. Graefes Arch Clin Exp Ophthalmol 229:457-63
Lin, W L (1990) Immunogold localization of basal laminar heparan sulfate proteoglycan in rat brain and retinal capillaries. Brain Res Bull 24:533-6
Lin, W L; Essner, E (1990) Immunogold localization of basement membrane molecules in rat retinal capillaries. Cell Mol Biol 36:13-21
Lin, W L (1989) Immunogold localization of extracellular matrix molecules in Bruch's membrane of the rat. Curr Eye Res 8:1171-8

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